scholarly journals Microinjection of antibodies against talin inhibits the spreading and migration of fibroblasts

1992 ◽  
Vol 102 (4) ◽  
pp. 753-762
Author(s):  
G.H. Nuckolls ◽  
L.H. Romer ◽  
K. Burridge
Keyword(s):  
Extracellular Matrix ◽  
Tissue Culture ◽  
Actin Filaments ◽  
Focal Adhesions ◽  
Cell Spreading ◽  
And Migration ◽  
Extracellular Matrix Receptors

Talin is believed to be one of the key proteins involved in linking actin filaments to extracellular matrix receptors in focal adhesions. Our strategy for studying the function of talin has been to inactivate talin in living fibroblasts in tissue culture through the microinjection of affinity-purified, polyclonal anti-talin antibodies. The effect of the injected anti-talin antibodies on cell spreading was found to depend on how recently the cells had been plated. Cells that were in the process of spreading on a fibronectin substratum, and which had newly developed focal adhesions, were induced to round up and to disassemble many of the adhesions. However, if fibroblasts were allowed to spread completely before they were microinjected with the anti-talin antibody, focal adhesions remained intact and the flat morphology of the cells was unaffected. The percentage of cells that were able to maintain a spread morphology despite the injection of anti-talin antibodies increased during the first few hours after plating on fibronectin substrata. Fibroblasts that were allowed to spread completely before microinjection with the anti-talin antibody retained both intact focal adhesions and a flat, well-spread morphology, but failed to migrate effectively. Our experiments do not directly address the role of talin in mature focal adhesions, but they indicate that talin is essential for the spreading and migration of fibroblasts on fibronectin as well as for the development and initial maintenance of focal adhesions on this substratum.

scholarly journals Syndecans and Pancreatic Ductal Adenocarcinoma

Biomolecules ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 349
Author(s):  
Nausika Betriu ◽  
Juan Bertran-Mas ◽  
Anna Andreeva ◽  
Carlos E. Semino
Keyword(s):  
Extracellular Matrix ◽  
Pancreatic Ductal Adenocarcinoma ◽  
Cell Types ◽  
Ductal Adenocarcinoma ◽  
Physiological Processes ◽  
Extracellular Matrix Components ◽  
Detection And Diagnosis ◽  
And Migration ◽  
Early Detection And Diagnosis

Pancreatic Ductal Adenocarcinoma (PDAC) is a fatal disease with poor prognosis because patients rarely express symptoms in initial stages, which prevents early detection and diagnosis. Syndecans, a subfamily of proteoglycans, are involved in many physiological processes including cell proliferation, adhesion, and migration. Syndecans are physiologically found in many cell types and their interactions with other macromolecules enhance many pathways. In particular, extracellular matrix components, growth factors, and integrins collect the majority of syndecans associations acting as biochemical, physical, and mechanical transducers. Syndecans are transmembrane glycoproteins, but occasionally their extracellular domain can be released from the cell surface by the action of matrix metalloproteinases, converting them into soluble molecules that are capable of binding distant molecules such as extracellular matrix (ECM) components, growth factor receptors, and integrins from other cells. In this review, we explore the role of syndecans in tumorigenesis as well as their potential as therapeutic targets. Finally, this work reviews the contribution of syndecan-1 and syndecan-2 in PDAC progression and illustrates its potential to be targeted in future treatments for this devastating disease.

scholarly journals Extracellular matrix plasticity as a driver of cell spreading

2020 ◽  
Vol 117 (42) ◽  
pp. 25999-26007
Author(s):  
Joshua M. Grolman ◽  
Philipp Weinand ◽  
David J. Mooney
Keyword(s):  
Extracellular Matrix ◽  
Stem Cell ◽  
Wound Repair ◽  
Kinetic Monte Carlo ◽  
Cell Spreading ◽  
Cell Biophysics ◽  
Biological Processes ◽  
Kinetic Monte Carlo Simulations ◽  
Network Plasticity ◽  
And Migration

Mammalian cell morphology has been linked to the viscoelastic properties of the adhesion substrate, which is particularly relevant in biological processes such as wound repair and embryonic development where cell spreading and migration are critical. Plastic deformation, degradation, and relaxation of stress are typically coupled in biomaterial systems used to explore these effects, making it unclear which variable drives cell behavior. Here we present a nondegradable polymer architecture that specifically decouples irreversible creep from stress relaxation and modulus. We demonstrate that network plasticity independently controls mesenchymal stem cell spreading through a biphasic relationship dependent on cell-intrinsic forces, and this relationship can be shifted by inhibiting actomyosin contractility. Kinetic Monte Carlo simulations also show strong correlation with experimental cell spreading data as a function of the extracellular matrix (ECM) plasticity. Furthermore, plasticity regulates many ECM adhesion and remodeling genes. Altogether, these findings confirm a key role for matrix plasticity in stem cell biophysics, and we anticipate this will have ramifications in the design of biomaterials to enhance therapeutic applications of stem cells.

scholarly journals Role of Actin Filaments in Correlating Nuclear Shape and Cell Spreading

PLoS ONE ◽  
2014 ◽  
Vol 9 (9) ◽  
pp. e107895 ◽  
Author(s):  
Renu Vishavkarma ◽  
Swetavalli Raghavan ◽  
Chandrashekar Kuyyamudi ◽  
Abhijit Majumder ◽  
Jyotsna Dhawan ◽  
...  
Keyword(s):  
Actin Filaments ◽  
Cell Spreading ◽  
Nuclear Shape

scholarly journals Tropomyosin Isoform Expression Regulates the Transition of Adhesions To Determine Cell Speed and Direction

2009 ◽  
Vol 29 (6) ◽  
pp. 1506-1514 ◽  
Author(s):  
Cuc T. T. Bach ◽  
Sarah Creed ◽  
Jessie Zhong ◽  
Maha Mahmassani ◽  
Galina Schevzov ◽  
...  
Keyword(s):  
Focal Adhesion ◽  
Actin Filaments ◽  
Feedback Loop ◽  
Cell Movement ◽  
Focal Adhesions ◽  
Leading Edge ◽  
Mesenchymal Cell ◽  
Critical Determinant ◽  
Focal Complex

ABSTRACT The balance of transition between distinct adhesion types contributes to the regulation of mesenchymal cell migration, and the characteristic association of adhesions with actin filaments led us to question the role of actin filament-associating proteins in the transition between adhesive states. Tropomyosin isoform association with actin filaments imparts distinct filament structures, and we have thus investigated the role for tropomyosins in determining the formation of distinct adhesion structures. Using combinations of overexpression, knockdown, and knockout approaches, we establish that Tm5NM1 preferentially stabilizes focal adhesions and drives the transition to fibrillar adhesions via stabilization of actin filaments. Moreover, our data suggest that the expression of Tm5NM1 is a critical determinant of paxillin phosphorylation, a signaling event that is necessary for focal adhesion disassembly. Thus, we propose that Tm5NM1 can regulate the feedback loop between focal adhesion disassembly and focal complex formation at the leading edge that is required for productive and directed cell movement.

scholarly journals Collagen IV regulates Caco-2 cell spreading and p130Cas phosphorylation by FAK-dependent and FAK-independent pathways

2008 ◽  
Vol 389 (1) ◽  
pp. 47-55 ◽  
Author(s):  
Matthew A. Sanders ◽  
Marc D. Basson
Keyword(s):  
Focal Adhesion ◽  
Cell Spreading ◽  
Spreading Rate ◽  
Collagen Iv ◽  
Intestinal Epithelial ◽  
Protein Levels ◽  
And Migration ◽  
Sirna Knockdown ◽  
Human Specific

Abstract We previously observed that collagen IV regulates Caco-2 intestinal epithelial cell spreading and migration via Src-dependent p130Cas phosphorylation and stimulates focal adhesion kinase (FAK). However, the role of FAK and the related kinase, Pyk2, in Caco-2 spreading and migration is unclear. FAK- or Pyk2-specific siRNAs reduced protein levels by 90%. However, when detached cells were replated on collagen IV neither individual nor combined FAK and Pyk2 siRNAs affected the cell spreading rate. As combined FAK and Pyk2 siRNAs increased p130Cas protein levels, we cotransfected cells with 1 nm p130Cas siRNA to partially reduce p130Cas protein to control levels. Although p130Cas Tyr(P)249 phosphorylation was reduced by 60%, cell spreading was unaffected. Combined siRNA reduction of FAK, Pyk2 and p130Cas increased cell spreading by 20% compared to p130Cas siRNA alone, suggesting that FAK and Pyk2 negatively regulate spreading in addition to stimulating spreading via p130Cas. FAK-binding mutant SH3 domain-deleted rat p130Cas was not phosphorylated after adhesion and, unlike full-length p130Cas, did not restore spreading after human-specific p130Cas siRNA knockdown of endogenous p130Cas. Together, these data suggest that FAK positively regulates Caco-2 spreading on collagen IV via p130Cas phosphorylation, but also suggests that FAK may negatively regulate spreading through other mechanisms and the presence of additional FAK-independent pathways regulating p130Cas.

scholarly journals LncRNA XIST promotes extracellular matrix synthesis, proliferation and migration by targeting miR-29b-3p/COL1A1 in human skin fibroblasts after thermal injury

2019 ◽  
Vol 52 (1) ◽  
Author(s):  
Wei Cao ◽  
Youping Feng
Keyword(s):  
Extracellular Matrix ◽  
Cell Proliferation ◽  
Wound Healing ◽  
Western Blot ◽  
Thermal Injury ◽  
Human Skin Fibroblasts ◽  
Cell Proliferation And Migration ◽  
And Migration ◽  
Proliferation And Migration

Abstract Background Long noncoding RNAs (lncRNAs) have been reported to be associated with dermis process during burn wound healing. This study aimed to investigate the role of lncRNA X-inactive specific transcript (XIST) in human skin fibroblasts (HSF) and extracellular matrix (ECM) as well as the regulatory network of XIST/microRNA-29b-3p (miR-29b-3p)/collagen 1 alpha 1 (COL1A1). Methods The wound samples were collected from 25 patients with deep partial thickness burn at day 5 after burn. The thermal injured model was established using HSF cells. The expressions of XIST, miR-29b-3p and COL1A1 were measured by quantitative real-time polymerase chain reaction and western blot. ECM synthesis, cell proliferation and migration were detected by western blot, cell counting kit-8 and trans-well assays, respectively. The interaction between miR-29b-3p and XIST or COL1A1 was explored by bioinformatics analysis and luciferase reporter assay. Results The expressions of XIST and COL1A1 were enhanced but miR-29b-3p expression was decreased after thermal injury. XIST overexpression promoted ECM synthesis, cell proliferation and migration in thermal injured HSF cells. However, XIST knockdown played an opposite effect. miR-29b-3p overexpression inhibited ECM synthesis, cell proliferation and migration, which was reversed by XIST. COL1A1 silence suppressed ECM synthesis, cell proliferation and migration by miR-29b-3p targeting. Moreover, COL1A1 up-regulation weakened the effect of XIST silence on ECM synthesis and HSF cell function. Conclusion XIST promoted ECM synthesis, cell proliferation and migration by sponging miR-29b-3p and targeting COL1A1 in HSF cells after thermal injury, indicating the promoting role of XIST in wound healing.

scholarly journals Focal adhesion kinase signaling is decreased in polyamine-depleted IEC-6 cells

2001 ◽  
Vol 281 (2) ◽  
pp. C475-C485 ◽  
Author(s):  
Ramesh M. Ray ◽  
Mary Jane Viar ◽  
Shirley A. McCormack ◽  
Leonard R. Johnson
Keyword(s):  
Extracellular Matrix ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Focal Adhesions ◽  
Stress Fiber ◽  
Fiber Formation ◽  
Integrin Signaling ◽  
Actin Stress Fiber ◽  
Stress Fiber Formation ◽  
And Migration

Polyamines are essential to the migration of epithelial cells in the intestinal mucosa. Cells depleted of polyamines do not attach as rapidly to the extracellular matrix and do not form the actin stress fibers essential for migration. Because both attachment and stress fiber formation depend on integrin signaling and the formation of focal adhesions, we examined these and related processes in polyamine-depleted IEC-6 cells. There was general decreased tyrosine phosphorylation of focal adhesion kinase (FAK), and, specifically, decreased phosphorylation of Tyr-925, the paxillin binding site. In control cells, FAK phosphorylation was rapid after attachment to the extracellular matrix, while attached cells depleted of polyamines had significantly delayed phosphorylation. FAK activity was also significantly inhibited in polyamine-depleted cells as was the phosphorylation of paxillin. Polyamine-depleted cells failed to spread normally after attachment, and immunocytochemistry showed little colocalization of FAK and actin compared with controls. Focal adhesion complex formation was greatly reduced in the absence of polyamines. These data suggest that defective integrin signaling may, at least in part, account for the decreased rates of attachment, actin stress fiber formation, spreading, and migration observed in polyamine-depleted cells.

The role of E-cadherin and integrins in mesoderm differentiation and migration at the mammalian primitive streak

Development ◽  
1993 ◽  
Vol 118 (3) ◽  
pp. 829-844 ◽  
Author(s):  
C.A. Burdsal ◽  
C.H. Damsky ◽  
R.A. Pedersen
Keyword(s):  
Extracellular Matrix ◽  
Primitive Streak ◽  
Intermediate Filament Protein ◽  
Causative Role ◽  
Adhesive Interactions ◽  
And Migration ◽  
E Cadherin ◽  
Cell Cell

We have examined the role of cell-cell and cell-extracellular matrix (ECM) interactions during mesoderm differentiation and migration at the primitive streak of the mouse embryo with the use of function-perturbing antibodies. Explants of epiblast or mesoderm tissue dissected from the primitive streak of 7.5- to 7.8-day mouse embryos were cultured on a fibronectin substratum in serum-free, chemically defined medium. After 16–24 hours in culture, cells in explants of epiblast exhibited the typical close-packed morphology of epithelia, and the tissue remained as a coherent patch of cells that were shown to express transcripts of the cytokeratin Endo B by in situ analysis. In contrast, cells in explants of primitive streak mesoderm exhibited a greatly flattened, fibroblastic morphology, did not express Endo B transcripts, and migrated away from the center of the explant. As epiblast cells in vivo undergo the epithelial-mesenchymal transition at the primitive streak, they cease expressing the prominent calcium-sensitive cell adhesion molecule E-cadherin (uvomorulin, Cell-CAM 120/80). We asked whether the loss of E-cadherin expression was a passive result of differentiation or if it might play a more causative role in mesoderm differentiation and migration. Culture with function-perturbing antibodies against E-cadherin caused cells within epiblast explants to lose cell-cell contacts, to flatten, and to assume a mesenchymal morphology; they were also induced to migrate. Anti-E-cadherin antibodies had no effect on explants of primitive streak mesoderm. In immunofluorescence studies, anti-E-cadherin-treated epiblast cells ceased to express SSEA-1, a carbohydrate moiety that is lost as mesoderm differentiates from the epiblast in vivo, and they also ceased to express E-cadherin itself. In contrast, these cells began to express the intermediate filament protein vimentin, a cytoskeletal protein characteristic of the primitive streak mesoderm at this stage of development. As epiblast cells differentiate into mesoderm, their predominant adhesive interactions change from cell-cell to cell-substratum. Therefore, we also investigated the adhesive interactions between primitive streak tissues and extracellular matrix (ECM) components. Epiblast explants adhered well to fibronectin, more poorly to laminin and type IV collagen, and not at all to vitronectin. In contrast, mesoderm explants attached well to all these proteins. Furthermore, epiblast, but not mesoderm, displayed an anchorage-dependent viability in culture. After anti-E-cadherin treatment, epiblast cells that had assumed the mesenchymal morphology did attach to vitronectin, another characteristic shared with primitive streak mesoderm.(ABSTRACT TRUNCATED AT 400 WORDS)

Smooth muscle cell rigidity and extracellular matrix organization influence endothelial cell spreading and adhesion formation in coculture

2007 ◽  
Vol 293 (3) ◽  
pp. H1978-H1986 ◽  
Author(s):  
Charles S. Wallace ◽  
Sophie A. Strike ◽  
George A. Truskey
Keyword(s):  
Extracellular Matrix ◽  
Smooth Muscle ◽  
Endothelial Cell ◽  
Focal Adhesion ◽  
Adhesion Formation ◽  
Focal Adhesions ◽  
Cell Spreading ◽  
Tissue Engineered Blood Vessels ◽  
Focal Adhesion Formation ◽  
Fibrillar Adhesion

Efforts to develop functional tissue-engineered blood vessels have focused on improving the strength and mechanical properties of the vessel wall, while the functional status of the endothelium within these vessels has received less attention. Endothelial cell (EC) function is influenced by interactions between its basal surface and the underlying extracellular matrix. In this study, we utilized a coculture model of a tissue-engineered blood vessel to evaluate EC attachment, spreading, and adhesion formation to the extracellular matrix on the surface of quiescent smooth muscle cells (SMCs). ECs attached to and spread on SMCs primarily through the α5β1-integrin complex, whereas ECs used either α5β1- or αvβ3-integrin to spread on fibronectin (FN) adsorbed to plastic. ECs in coculture lacked focal adhesions, but EC α5β1-integrin bound to fibrillar FN on the SMC surface, promoting rapid fibrillar adhesion formation. As assessed by both Western blot analysis and quantitative real-time RT-PCR, coculture suppressed the expression of focal adhesion proteins and mRNA, whereas tensin protein and mRNA expression were elevated. When attached to polyacrylamide gels with similar elastic moduli as SMCs, focal adhesion formation and the rate of cell spreading increased relative to ECs in coculture. Thus, the elastic properties are only one factor contributing to EC spreading and focal adhesion formation in coculture. The results suggest that the softness of the SMCs and the fibrillar organization of FN inhibit focal adhesions and reduce cell spreading while promoting fibrillar adhesion formation. These changes in the type of adhesions may alter EC signaling pathways in tissue-engineered blood vessels.

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